PROJECT SUMMARY Billions of people are at risk of contracting vector-borne diseases and there are no drugs or vaccines to treat or prevent the majority of these infections. Therefore, vector control is the primary tool used for vector-borne disease prevention. This proposal aims to improve the technologies available to conduct functional analysis of genes in ?Ae. aegypti?, a major vector of dengue, Zika, chikungunya and yellow fever. Additionally, these same tools can be adapted to develop antiviral effectors for genetic population replacement strategies, which could be a key tool for the sustainable management of vector-borne disease. While the goal of creating functional analytic tools for mosquitoes is not novel, the proposed RNA guided, RNA targeting endonuclease tools will likely be a more efficient technology with less off target effects compared to current RNAi screening technologies. Furthermore, we use these new technologies to develop antiviral effectors for the development of disease refractory mosquitoes, which will expect to be an efficient alternative to current RNAi-based antiviral effectors. Therefore, this project will adapt RNA guided, RNA targeting Cas endonucleases to develop functional transcriptomic tools and novel post-transcriptional and viral RNA silencing strategies in ?Ae. aegypti?. The tools will be first developed in mosquito cells and then transitioned into an ?in vivo ?mosquito model. Comparative analysis of the efficiency of RNA guided, RNA targeting Cas endonuclease tools, RNAi and Cas9 functional ?omics? tools will also be conducted to determine the most robust RNA silencing and functional genomic analytical strategies. Finally, these tools will be designed to target viral RNA in mosquito cells and in the whole organism. In the end, we will develop a functional transcriptomics tool that will facilitate the characterization of ?Ae. aegypti ?genes as well as the development of novel antiviral effectors.